CN113322291A

CN113322291A - Synthesis method of chiral amino alcohol compound

Info

Publication number: CN113322291A
Application number: CN202010127746.4A
Authority: CN
Inventors: 彭要武; 田文敬; 叶青; 胡磊; 方志成
Original assignee: Masteam Bio Tech Co ltd
Current assignee: Masteam Bio Tech Co ltd
Priority date: 2020-02-28
Filing date: 2020-02-28
Publication date: 2021-08-31

Abstract

The invention discloses a method for synthesizing a chiral amino alcohol compound. The synthesis method of the chiral amino alcohol compound provided by the invention comprises the following steps: in an organic solvent and a buffer solution, under the existence of carbonyl reductase and coenzyme, carrying out the reduction reaction shown as the following on a carbonyl compound shown as a formula II to obtain a conversion solution containing a chiral amino alcohol compound shown as a formula I; the carbonyl reductase is carbonyl reductase from Rhodotorula toruloides (Rhodotorula toruloides); wherein R is¹Is (S) -1-phenylethyl and/or (R) -1-phenylethyl. According to the invention, racemic azapiperidine chiral compound is used as a starting material, and the chiral amino alcohol with a single configuration is efficiently synthesized through an enzymatic reaction of recombinant carbonyl reductase, so that an important intermediate with a high de value is obtained, and the production cost is favorably reduced.

Description

Synthesis method of chiral amino alcohol compound

Technical Field

The invention relates to a method for synthesizing a chiral amino alcohol compound.

Background

Florfenicol (Florfenicol) is a special broad-spectrum antibiotic of chloramphenical for animals, which is developed by Nagab-hushan and the like of Proben-Baoya (Schering-Plough) in the 20 th century at the end of 70 years. In view of the fact that the drug effect of florfenicol is superior to that of chloramphenicol and thiamphenicol in the prevention and treatment of animal diseases, the florfenicol has a wider application prospect, and the synthesis of the florfenicol is always paid great attention.

In recent years, the research of synthesizing chiral compounds by using enzymatic reduction is relatively extensive, two Chinese patents, namely CN102827042A and CN103936638A, are invented in the florfenicol new process development process by the company, wherein in the patent CN103936638A, a very important intermediate chiral amino alcohol with a single configuration is synthesized by a chemical method and needs chemical resolution and chiral reduction. And the efficiency is low.

The synthetic route in patent CN103936638A is as follows:

the patent CN106316898B discloses the synthetic route as follows:

the method obtains chiral amino alcohols with different configurations through different reducing reagents, chiral aminocarbonyl is obtained through chemical resolution, and a chiral aminocarbonyl compound with a single configuration is obtained through repeated recrystallization.

Disclosure of Invention

The invention aims to solve the problem of overcoming the defect of single preparation method of chiral amino alcohol of a key intermediate of florfenicol in the prior art, and provides a biosynthesis method of a chiral amino alcohol compound; the method is characterized in that a single chiral amino alcohol compound is prepared by a microbial/enzyme catalysis technology, an achiral precursor and a biological catalysis synthesis system through a biological catalysis and conversion method. The biocatalytic synthesis system used by the method has the characteristics of high efficiency, low cost and no pollution; has high efficiency, high yield, low cost and high commercial application value.

The present invention solves the above-mentioned problems by the following technical means.

The invention provides a method for synthesizing a chiral amino alcohol compound, which comprises the following steps: in an organic solvent and a buffer solution, under the existence of carbonyl reductase and a coenzyme system, carrying out the reduction reaction shown as the following on a carbonyl compound shown as a formula II to obtain a conversion solution containing a chiral amino alcohol compound shown as a formula I; the carbonyl reductase is carbonyl reductase from Rhodotorula toruloides (Rhodotorula toruloides);

wherein R is¹Is (S) -1-phenylethyl and/or (R) -1-phenylethyl;

represents a chiral carbon, which is in S configuration or a mixture of S and R configurations.

In one embodiment of the present invention, the carbonyl reductase (CGMCC 2.1389) derived from Rhodotorula toruloides is selected from the group consisting of:

(i) the amino acid sequence is shown as SEQ ID NO. 1;

and/or, (ii) substitution, deletion, alteration, insertion or addition of one or more amino acids of the amino acid sequence shown in SEQ ID NO.1 within a range that retains the activity of the enzyme, and the resulting amino acid sequence, for example, has a homology of 95% or more with the amino acid sequence shown in SEQ ID NO. 1.

In one embodiment of the present invention, the gene sequence encoding carbonyl reductase is selected from the group consisting of:

(a) a sequence shown as SEQ ID NO. 2;

(b) a polynucleotide complementary to the sequence defined in (a); or

(c) Any polynucleotide or complementary sequence having at least 70%, preferably at least 75%, 80%, 85%, 90%, more preferably at least 95%, 96%, 97%, 98%, 99% or more sequence identity to the sequence defined in (a).

In one embodiment of the present invention, the carbonyl reductase may be in a form conventional in the art, such as free form enzyme, lyophilized powder, broken enzyme solution, immobilized enzyme, or enzyme in bacterial form (carbonyl reductase genetically engineered bacteria whole cell); the invention is preferably a gene engineering bacterium whole cell disruption enzyme solution containing the carbonyl reductase.

In one aspect of the present invention, when the carbonyl reductase is a whole cell disruption enzyme solution of a genetically engineered bacterium containing the carbonyl reductase, the mass ratio of the carbonyl compound represented by the formula II to the enzyme solution may be 2: 1.

In one aspect of the present invention, in the reduction reaction, when the carbonyl reductase is a whole cell disruption enzyme solution of a genetically engineered bacterium containing the carbonyl reductase, the concentration of the whole cell disruption enzyme solution of the genetically engineered bacterium may be 50g/L to 100g/L (e.g., 50 g/L); the enzyme activity of the carbonyl reductase can be 100 mu/mg.

In one embodiment of the present invention, the carbonyl reductase is preferably (R, R) -carbonyl reductase.

The organic solvent may be an organic solvent which is conventional in such reactions in the art, and in the present invention, one or more of DMSO, DMF, 3 ', 5, 5' -tetramethylbiphenyl dihydroxyethyl ether (TMBE), and methanol are preferable. The organic solvent may be used in an amount which is conventional in such reactions and does not interfere with the reaction, for example, to dissolve the substrate. In one aspect of the present invention, the concentration of the organic solvent in the reduction reaction is from 10% by volume (for example, 5%) to 10% by volume of the reaction system.

In the present invention, the buffer solution may be a buffer solution conventional in such reactions in the art, such as phosphate buffer solution, carbonate buffer solution, Tri-HCl buffer solution, borate buffer solution, glycine buffer solution, citrate buffer solution, or MOPS buffer solution; in one embodiment of the present invention, a Tris-hydrochloric acid buffer solution (Tris: Tris (hydroxymethyl) aminomethane, for example, at a concentration of 0.05mol/L, 25 ℃ C.) is preferred.

In one embodiment of the present invention, the pH of the buffer solution may be a pH conventional in such reactions in the art, such as 7.0 to 9.0; preferably the pH is 8.0.

In one embodiment of the present invention, the molar percentage of the coenzyme in the coenzyme system to the carbonyl compound represented by formula II may be 0.01% to 1.0%, preferably 0.1% to 0.6%.

In a certain aspect of the present invention, in the reduction reaction, the concentration of the coenzyme in the coenzyme system in the reaction system may be 1mM-10mM (e.g., 5 mM).

In one embodiment of the present invention, the coenzyme in the coenzyme system is selected from the group consisting of: a reducing coenzyme, an oxidizing coenzyme, or a combination thereof; for example NADH, NADPH, NAD⁺、NADP⁺Or a combination thereof; preferably NADH, NAD⁺Or a combination thereof. When the coenzyme in the coenzyme system contains NAD⁺When used, the enzyme (dehydrogenase) for the coenzyme regeneration and the corresponding cosubstrate (hydrogen donor) may be included; the regenerated enzyme may be selected from the group consisting of: an alcohol dehydrogenase, a formate dehydrogenase, a glucose dehydrogenase, or a combination thereof; the co-substrate is selected from the group consisting of: isopropanol, glucose, ammonium formate, or a combination thereof; preferably, the regenerated enzyme and its corresponding co-substrate are glucose dehydrogenase (EC1.1.1.47) and glucose.

In one embodiment of the present invention, when the coenzyme contains NAD⁺When the coenzyme comprises a regenerated enzyme and its corresponding co-substrate, the molar ratio of the co-substrate to the carbonyl compound of formula II may be a molar ratio as is conventional in reactions of this type in the art, e.g.1.5: 1.

In one embodiment of the present invention, the degree of co-substrate in the reduction reaction may be 90 g/L.

In one aspect of the present invention, in the reduction reaction, when the coenzyme in the coenzyme system contains NAD⁺The concentration of the regenerated enzyme in the reaction system when the regenerated enzyme and the corresponding co-substrate thereof are used can be 25 mg/L; the regenerated enzyme may have an enzyme activity of 500. mu.g/mg.

In one embodiment of the invention, the temperature of the reduction reaction may be a temperature conventional in such reactions in the art, such as 10 ℃ to 50 ℃, preferably 20 ℃ to 40 ℃, and more preferably 25 ℃ to 35 ℃.

In one embodiment of the present invention, the time for the reduction reaction may be 1 to 120 hours, preferably 5 to 72 hours, and more preferably 15 to 24 hours.

In a certain embodiment of the present invention, the method for synthesizing chiral amino alcohol compounds comprises the following steps: controlling the pH value to be 8.0 in an organic solvent and a Tris-hydrochloric acid buffer solution, and carrying out the reduction reaction on a carbonyl compound shown as a formula II in the presence of a carbonyl reductase and a coenzyme system to obtain a conversion solution containing a chiral amino alcohol compound shown as a formula I; the carbonyl reductase is carbonyl reductase from rhodotorula toruloides; the coenzyme system comprises an oxidative coenzyme NAD⁺The enzyme for coenzyme regeneration is glucose dehydrogenase and its corresponding co-substrate is glucose. Preferably, the amino acid sequence of the carbonyl reductase is shown as SEQ ID NO.1, and/or the amino acid sequence with homology of more than 95% with the amino acid sequence shown as SEQ ID NO. 1.

In one embodiment of the present invention, the synthesis method further comprises: and after the reduction reaction is finished, separating the chiral amino alcohol compound shown in the formula I from the conversion solution. The isolation step may be one conventional in such reactions in the art and may comprise: centrifuging the cells, adding 5% NaOH to the supernatant, filtering to remove precipitated solid, extracting the remaining conversion solution with an organic solvent (e.g., dichloromethane), washing (e.g., water or saturated saline), drying (e.g., conventional anhydrous sodium sulfate), and concentrating; obtaining the chiral amino alcohol compound shown in the formula I. The organic solvent may be an organic solvent conventional in such extractions in the art, such as methylene chloride.

In a certain scheme of the invention, the de value of the chiral amino alcohol compound shown in the formula I is more than or equal to 90%, preferably more than or equal to 95%, and more preferably more than or equal to 99%.

In one embodiment of the present invention, the conversion rate of the carbonyl compound shown in formula II to the chiral amino alcohol compound shown in formula I is greater than or equal to 80%, such as greater than or equal to 85%, preferably greater than or equal to 95%, and more preferably greater than or equal to 99%.

The present invention also provides a reaction system comprising:

organic solvent, buffer solution, carbonyl reductase, coenzyme, enzyme for coenzyme regeneration and corresponding cosubstrate thereof, and carbonyl compound shown in formula II;

wherein the organic solvent, the buffer solution, the carbonyl reductase, the coenzyme, the enzyme for coenzyme regeneration and the corresponding co-substrate thereof, the carbonyl compound represented by formula II, are as defined above.

In one aspect of the present invention, the reaction system consists of:

organic solvent, buffer solution, carbonyl reductase, coenzyme, enzyme for coenzyme regeneration and corresponding cosubstrate thereof, and carbonyl compound shown as formula II.

Term(s) for

Enantiomeric excess (ee, enantiomeric excess): are commonly used to characterize the excess of one enantiomer relative to the other in chiral molecules.

Diastereomeric excess (de, diasteromeric processes): are commonly used to characterize the excess of one diastereomer over another in molecules with two or more chiral centers.

In the present invention, "carbonyl reductase" refers to an enzyme capable of stereoselectively asymmetrically catalytically reducing a prochiral ketone to a chiral alcohol. Stereoselectivity is defined as enantiomeric excess (ee) of 80% or more and diastereomeric excess (de) of 80% or more.

In the invention, the configuration is defined by taking the chiral amino alcohol compound shown as the formula I as a reference, wherein the configuration of the hydroxyl is R, the configuration of the chiral carbon on the aziridine is R, and any carbonyl reductase which can stereoselectively identify the chiral carbon on the aziridine with the R-configuration in the carbonyl compound shown as the formula II and reduce the carbonyl in the carbonyl compound shown as the formula II into the hydroxyl with the R-configuration is defined as (R, R) -carbonyl reductase in the technical scheme of the invention.

In the present invention, the carbonyl reductase may be wild-type or mutant. Furthermore, they may be isolated or recombinant.

Carbonyl reductases useful in the present invention may be from different species. The amino acid sequence of a typical carbonyl reductase is shown as SEQ ID No.1, and the coding gene is shown as SEQ ID No. 2.

SEQ ID No.1

SEQ ID No.2

The carbonyl reductase of the invention also comprises an amino acid sequence obtained by replacing, deleting, changing, inserting or adding one or more amino acids of the amino acid sequence shown in SEQ ID NO.1 within the range of keeping the activity of the enzyme.

In the present invention, the carbonyl reductase may be used in various forms. For example, resting cells or wet cells expressing the carbonyl reductase of the present invention may be used, various forms such as crude enzyme solution, pure enzyme or crude enzyme powder may be used, or immobilized enzyme may be used. Preferably, in order to obtain higher conversion efficiency and reduce cost, a crude enzyme solution, such as a whole cell disruption enzyme solution of a genetically engineered bacterium containing the carbonyl reductase, is preferably used.

In the invention"coenzyme" means a coenzyme capable of effecting electron transfer in a redox reaction. The coenzyme of the invention is a reductive coenzyme NADH, NADPH or an oxidative coenzyme NAD⁺、NADP⁺. Since the reducing coenzyme is expensive, the oxidizing coenzyme NAD is preferred⁺、NADP⁺. When the oxidative coenzyme is selected, a method for realizing coenzyme regeneration needs to be selected, and the method mainly comprises three types of (1) glucose dehydrogenase and cosubstrate glucose; (2) alcohol dehydrogenase and co-substrate isopropanol; (3) formate dehydrogenase co-substrate ammonium formate. In a preferred embodiment, the coenzyme comprises NAD⁺The coenzyme regeneration system is glucose dehydrogenase, oxidative coenzyme NAD⁺The mol percentage of the dosage to the dosage of the substrate is 0.01 percent to 1.0 percent, and the buffer system is 0.05mol/L Tris-hydrochloric acid buffer solution. The pH of the buffer solution is 7.0-9.0.

In the present invention, a co-solvent may be added or not added to the reaction system. The term "co-solvent" refers to a sparingly soluble substance that forms a soluble intermolecular complex, association, double salt, or the like with an added third substance in a solvent to increase the solubility of the sparingly soluble substance in the solvent. This third material is referred to as a co-solvent.

In the present invention, unless otherwise specified, the concentration is the final concentration of the compound in the whole reaction system before the reaction.

The above preferred conditions can be arbitrarily combined to obtain preferred embodiments of the present invention without departing from the common general knowledge in the art.

The carbonyl reductase and the 1-phenethyl-2- [4- (methylsulfonyl) phenyl ] formacyl aziridine are self-made, and other used reagents and raw materials are commercially available.

The positive progress effects of the invention are as follows: the chiral amino alcohol which is an important intermediate in the novel process for preparing the florfenicol by using the enzyme method to carry out the asymmetric reduction reaction avoids chemical resolution and chiral reduction, and the required chiral amino alcohol compound is directly obtained through the specificity and specificity of the enzyme, so the process steps can be shortened, and the industrial production is facilitated. The method has the advantages of strong stereoselectivity, mild reaction conditions, simple and convenient operation, low cost, less pollution, less catalyst consumption, high efficiency, yield of more than 90 percent and high optical purity of the product (chiral de value of more than 99.5 percent); and two chiral centers are constructed by one-step reaction, so that the production efficiency is greatly improved, and the production cost is reduced.

Detailed Description

The invention is further illustrated by the following examples, which are not intended to limit the scope of the invention. The experimental procedures, for which specific conditions are not indicated in the following examples, are selected according to conventional methods and conditions (e.g., conditions described in Sambrook et al, molecular cloning: A Laboratory Manual (New York: Cold Spring Harbor Laboratory Press, 1989)), or according to commercial instructions.

Carbonyl reductase and 1-phenethyl-2- [4- (methylsulfonyl) phenyl ] formyl aziridine in the following examples are self-made, which are merely exemplary, and other sources are also suitable for use in the present invention as long as they meet the requirements of carbonyl reductase, coenzyme and 1-phenethyl-2- [4- (methylsulfonyl) phenyl ] formyl aziridine.

The experimental procedures in the present invention are conventional unless otherwise specified.

The HPLC analysis method comprises the following steps: a chromatographic column: asahi ultimate LP-C18(4.6 × 250mm,5 μm); column temperature: 35 ℃; flow rate: 0.8 mL/min; detection wavelength: 224 nm; the mobile phase is acetonitrile: 50mM potassium dihydrogen phosphate buffer (pH 5.5) 4: 6. Isocratic elution.

In the present invention, the construction of the whole cell of the genetically engineered bacteria of carbonyl reductase can be carried out by conventional methods in the art, for example, the following steps:

1. optimization of carbonyl reductase genes

The basic method is to eliminate some low-usage codons by optimizing the codons of carbonyl reductase according to the whole genome sequence of Rhodotorula toruloides, and simultaneously optimize the primary structure of mRNA by synonymy transformation, so that ribosome can smoothly translate backwards along the initiation codon. And eliminating Nde I and Xho I cleavage sites by using a synonymous transformation method. Synthesizing carbonyl reductase gene through whole gene synthesis;

2. construction of carbonyl reductase Gene expression vector

The basic method is to synthesize an optimized carbonyl reductase gene by a whole-gene synthesis method, and then connect a carbonyl reductase gene fragment to a pET-28a plasmid according to a designed enzyme cutting site to construct a recombinant plasmid;

3. construction of carbonyl reductase recombinant expression Strain

The basic method is to transform the constructed recombinant plasmid into Escherichia coli, thereby obtaining the recombinant Escherichia coli which over-expresses carbonyl reductase, namely engineering bacteria whole cells of the carbonyl reductase.

Example 1

The step (1) is a gene engineering bacterium of carbonyl reductase from Rhodotorula toruloides, and the specific preparation method is as follows:

(1) constructing an expression vector containing a carbonyl reductase gene:

rhodotorula toruloides (Rhodotorula torula)toruloides is purchased from China microorganism culture collection center (preservation number CGMCC 2.1389), carbonyl reductase from rhodotorula toruloides is subjected to random mutation, strains with higher enzyme activity are screened, codon GC content and rare codons are optimized, and whole gene synthesis is carried out to obtain carbonyl reductase, so as to obtain pET-28a recombinant plasmid. The optimized amino acid sequence is shown as SEQ ID No. 1.

(2) Transformation of the recombinant plasmid into a host cell

The recombinant plasmid was transformed into competent Escherichia coli BL21(DE3) (Tiangen Biochemical technology Co., Ltd.), spread on LB solid medium containing 50. mu.g/mL kanamycin, and cultured at 37 ℃ for 20 to 24 hours to obtain a primary positive clone.

(3) Screening by resistance culture medium to obtain positive clone

Respectively picking the primary positive clones in 5mL LB liquid culture medium containing 50 mug/mL kanamycin, culturing overnight at 37 ℃ and 200rpm, extracting plasmids, carrying out restriction enzyme NdeI and XhoI double enzyme digestion on the plasmids, carrying out electrophoresis verification, sequencing the extracted plasmids, and after verifying that no errors exist, determining the colony with the plasmids as the positive clone, namely the carbonyl reductase gene engineering bacteria derived from rhodotorula toruloides.

(4) The expression method of carbonyl reductase comprises the following steps: inoculating carbonyl reductase gene engineering bacteria into LB liquid culture medium added with 50 mug/mL kanamycin, and carrying out shake culture at 37 ℃ overnight; then transferred to LB liquid medium containing 50. mu.g/mL kanamycin in an inoculum size of 2% (v/v) and cultured to OD₆₀₀And (3) after the concentration is 0.8, adding IPTG (isopropyl-beta-D-thiogalactoside) to perform induction, wherein the final concentration of IPTG is 0.2mmol/L, cooling to 25 ℃, performing induction for 16h, and centrifuging to collect thalli to obtain the whole cell of the carbonyl reductase gene engineering bacteria derived from rhodotorula toruloides.

And carrying out ultrasonic crushing on the obtained whole cell of the genetically engineered bacteria with the carbonyl reductase activity by adopting an ultrasonic crushing method to obtain a carbonyl reductase genetically engineered bacteria whole cell crushed enzyme solution.

The glucose dehydrogenase used was a commercial enzyme from sigma-aldrich (CAS: 9028-53-9).

Step (2) enzymatic conversion reaction

The reaction is carried out in a 1L shake flask, the reaction system is controlled to be 300mL, and 1-phenethyl-2- [4- (methylsulfonyl) phenyl group]Formyl aziridine as a substrate (33.0g, 0.1mol) was dissolved in 15mL of DMSO, glucose (27.0g, 0.15mol) was added, a Tris-HCl buffer solution pH 8.0 was used as a solvent, whole cells of a genetically engineered bacterium derived from a carbonyl reductase derived from Rhodotorula torula, and glucose dehydrogenase were used as catalysts (500. mu.g/mg, 25mg/L), and coenzyme NAD was added⁺. Controlling carbonyl reductase derived from Rhodotorula toruloides (100 μ/mg. measuring the enzymatic activity of reductase pure protein, reaction system is 0.25ml, comprises Tris-HCl, pH 8.0, 2mmol/L NADH, 0.1 mmol/L1-phenethyl-2- [4- (methylsulfonyl) phenyl group]A formyl aziridine compound, and an amount of an enzyme, measured as a decrease in absorbance at 340nm, with the enzyme activity unit (U) defined as: the amount of enzyme required to catalyze the oxidation of 1. mu. mol NADH per minute under the above conditions. ) The whole cell concentration of the genetic engineering bacteria is 50g/L, and coenzyme NAD is controlled⁺Is 5 mM. Controlling the pH value of the conversion system to be 8.0, the conversion temperature to be 30 ℃, and controlling the rotating speed of a shaking table to be160r/min (monitored by HPLC until disappearance of starting material) the conversion time was 18 h. After conversion, the (R) - [4- (methylsulfonyl) phenyl is obtained][ (R) -1-phenylethyl-aziridin-2-yl]A methanol conversion solution.

Step (3) preparation of product

Purifying the conversion solution containing the compound (R) - [4- (methylsulfonyl) phenyl ] [ (R) -1-phenethyl-aziridin-2-yl ] methanol obtained in step (2), comprising the following steps: and centrifuging to collect cells, adding 5% NaOH into the obtained supernatant, stirring for half an hour to obtain a large amount of white solid, filtering, extracting the reaction solution with dichloromethane for 2 times, combining organic layers, washing with water once, washing with saturated salt water once, drying with anhydrous sodium sulfate, and spin-drying to obtain 29.7g of a product. The conversion was 99.3%, giving (R) - [4- (methylsulfonyl) phenyl ] [ (R) -1-R1-aziridin-2-yl ] methanol in an ee of 99.8%, DE of 99.5% and a yield of 90%. Wherein the target product [4- (methylsulfonyl) phenyl ] [ (R) -1-phenethyl-aziridin-2-yl ] methanol has a retention time of 6.90, and the target product [4- (methylsulfonyl) phenyl ] [ (S-1-phenethyl-aziridin-2-yl ] methanol has a retention time of 4.76.)

SEQUENCE LISTING

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Claims

1. A method for synthesizing chiral amino alcohol compounds is characterized by comprising the following steps: in an organic solvent and a buffer solution, under the existence of carbonyl reductase and coenzyme, carrying out the reduction reaction shown as the following on a carbonyl compound shown as a formula II to obtain a conversion solution containing a chiral amino alcohol compound shown as a formula I; the carbonyl reductase is carbonyl reductase from Rhodotorula toruloides (Rhodotorula toruloides);

wherein R is¹Is (S) -1-phenylethyl and/or (R) -1-phenylethyl;

2. The method of synthesis according to claim 1,

the carbonyl reductase is selected from the following (i) and/or (ii):

(i) the amino acid sequence is shown as SEQ ID NO. 1;

(ii) the amino acid sequence obtained by replacing, deleting, changing, inserting or adding one or more amino acids of the amino acid sequence shown in SEQ ID NO.1 in the range of keeping the enzyme activity;

and/or the carbonyl reductase is free enzyme, freeze-dried powder, broken enzyme liquid, immobilized enzyme or enzyme in the form of thalli.

3. The method of synthesis according to claim 2,

the coding gene sequence of the carbonyl reductase is selected from the following group:

(a) a sequence shown as SEQ ID NO. 2;

(b) a polynucleotide complementary to the sequence defined in (a); or

(c) Any polynucleotide or complementary sequence having at least 70% or more sequence identity to a sequence defined in (a);

and/or the carbonyl reductase is a gene engineering bacterium whole cell disruption enzyme liquid containing the carbonyl reductase.

4. The method of synthesis according to claim 3,

when the carbonyl reductase is a gene engineering bacterium whole-cell broken enzyme solution containing the carbonyl reductase, the mass ratio of the carbonyl compound shown as the formula II to the enzyme solution is 2: 1;

and/or, when the carbonyl reductase is a gene engineering bacterium whole-cell broken enzyme solution containing the carbonyl reductase, the concentration of the gene engineering bacterium whole-cell broken enzyme solution is 50g/L-100 g/L;

and/or the enzyme activity of the carbonyl reductase is 100 mu/mg;

and/or the carbonyl reductase is (R, R) -carbonyl reductase.

5. The synthetic method of any one of claims 1-4 wherein,

the organic solvent is one or more of DMSO, DMF, TMBE and methanol;

and/or the volume percentage concentration of the organic solvent is 1-10 percent;

and/or the buffer solution is phosphate buffer solution, carbonate buffer solution, Tri-HCl buffer solution, borate buffer solution, glycine buffer solution, citrate buffer solution or MOPS buffer solution;

and/or the pH value of the buffer solution is 7.0-9.0;

and/or, the coenzyme is NADH, NADPH, NAD⁺、NADP⁺Or a combination thereof;

and/or the molar percentage of the coenzyme to the carbonyl compound shown in the formula II is 0.01-1.0%;

and/or, the concentration of the coenzyme is 1mM-10 mM;

and/or the temperature of the reduction reaction is 10-50 ℃.

6. The method of synthesis according to claim 5,

the organic solvent is DMSO;

and/or the volume percentage concentration of the organic solvent is 5 percent;

and/or the buffer solution is Tris-hydrochloric acid buffer solution;

and/or the pH value of the buffer solution is 8.0;

and/or, the coenzyme is NADH, NAD⁺Or a combination thereof;

and/or the molar percentage of the coenzyme to the carbonyl compound shown in the formula II is 0.1-0.6%;

and/or the concentration of the coenzyme is 5 mM;

and/or, when the coenzyme in the coenzyme system comprises NAD⁺Said coenzyme system further comprising an enzyme for regeneration of said coenzyme and its corresponding co-substrate;

and/or the temperature of the reduction reaction is 20-40 ℃.

7. The method of synthesis according to claim 6,

the coenzyme is NAD⁺；

And/or, when the coenzyme in the coenzyme system comprises NAD⁺Regenerated enzyme and its corresponding co-substrate, said regenerated enzyme is selected from the group consisting of: an alcohol dehydrogenase, a formate dehydrogenase, a glucose dehydrogenase, or a combination thereof;

and/or, when the coenzyme in the coenzyme system comprises NAD⁺Regenerated enzyme and its corresponding co-substrate, said co-substrate being selected from the group consisting of: isopropanol, glucose, ammonium formate, or a combination thereof;

and/or, when the coenzyme bodyThe coenzyme in the system contains NAD⁺The regenerated enzyme and the corresponding co-substrate thereof, wherein the molar ratio of the co-substrate to the carbonyl compound shown in the formula II is 1.5: 1;

and/or the temperature of the reduction reaction is 25-35 ℃.

8. The method of synthesis according to claim 7,

when the coenzyme in the coenzyme system contains NAD⁺The regenerated enzyme and its corresponding co-substrate are glucose dehydrogenase and glucose;

and/or, when the coenzyme in the coenzyme system comprises NAD⁺Regenerated enzyme and its corresponding cosubstrate, the concentration of said regenerated enzyme is 25 mg/L; the enzyme activity of the regenerated enzyme is 500 mu/mg;

and/or, when the coenzyme in the coenzyme system comprises NAD⁺Regenerated enzyme and its corresponding co-substrate, the concentration of said co-substrate is 90 g/L.

9. The synthetic method of any one of claims 1-4 wherein,

the synthesis method of the chiral amino alcohol compound comprises the following steps: controlling the pH value to be 8.0 in an organic solvent and a Tris-hydrochloric acid buffer solution, and carrying out the reduction reaction on a carbonyl compound shown as a formula II in the presence of a carbonyl reductase and a coenzyme system to obtain a conversion solution containing a chiral amino alcohol compound shown as a formula I; the carbonyl reductase is carbonyl reductase from rhodotorula toruloides; the coenzyme system comprises an oxidative coenzyme NAD⁺The enzyme for coenzyme regeneration is glucose dehydrogenase and the corresponding co-substrate is glucose;

and/or, the synthesis method further comprises post-treatment, wherein the post-treatment comprises the following steps: after the reduction reaction is finished, centrifuging the thalli, adding 5% NaOH into the obtained supernatant, filtering to remove precipitated solids, extracting the residual conversion solution by using an organic solvent, washing, drying and concentrating; obtaining the chiral amino alcohol compound shown in the formula I.

10. A reaction system, characterized in that the reaction system comprises:

wherein the organic solvent, the buffer solution, the carbonyl reductase, the coenzyme, the enzyme for coenzyme regeneration and the corresponding co-substrate thereof, the carbonyl compound of formula II are as defined in any one of claims 1 to 9.